大豆食叶性害虫是影响大豆产量和品质的因素之一，本研究采用室内喂养斜纹夜蛾的方法，利用网室内采集的58份大豆资源的叶片，室内喂养相同龄期的斜纹夜蛾幼虫。根据虫体的反应，以幼虫重和蛹重为指标，研究了大豆对斜纹夜蛾单一虫种的抗性。综合2004和2005年的鉴定结果，获得了稳定性较好的高抗品种（黄皮小青豆、山东大豆、PI229358）和高感品种（徐疃大豆、沔阳白毛豆、枞阳猴子毛）。为克隆抗虫基因、鉴定基因功能以及研究大豆对斜纹夜蛾抗生性的遗传提供稳定可靠的抗、感材料。 本研究利用RACE的方法从大豆中克隆了茉莉酸生物合成的关键酶GmAOS基因及其启动子。GmAOS基因全长1789 bp，包含完整的ORF，编码519个氨基酸，没有内含子，GmAOS基因序列上有多个丝氨酸、苏氨酸、酪氨酸的磷酸化位点，GmAOS基因在大豆不同抗性的材料中均有2个拷贝数；软件分析显示，GmAOS基因蛋白质等电点8.97，分子量58.3 KDa，具有典型叶绿体定位信号肽，其二级结构主要由β折叠和α螺旋构成。GmAOS启动子区有2个TATA-box、1个CAAT-box、1个响应赤霉素的作用元件（TAACAA）、1个响应可诱导性抗性基因的作用元件（W box），2个响应细菌和盐诱导的作用元件（GAAAAA）和1个茉莉酸诱导表达所必须的顺式元件（G box）。 利用对斜纹夜蛾表现高抗、高感的大豆材料，检测了GmAOS基因及其同源基因GmAOS1（ABB917763）在不同抗性材料中组织表达量的差异和不同胁迫诱导表达量的差异。 GmAOS基因和GmAOS1基因在黄皮小青豆（高抗）和徐疃大豆（高感）的根、茎、叶、茎尖中都有表达，但是GmAOS基因和GmAOS1基因在黄皮小青豆（高抗）中的表达量比在徐疃大豆（高感）中的表达量高。GmAOS基因和GmAOS1基因对非生物胁迫冷(4℃)/热(45℃)/JA/SA/CuSO4<下标!>/Cut和生物胁迫SMV/Cotton worm均有响应；斜纹夜蛾诱导时，GmAOS 和GmAOS1基因达到最大表达量在黄皮小青豆（高抗）中需要3小时，而GmAOS基因达到最大表达量在徐疃大豆（高感）中需要24小时，GmAOS1基因需要12小时。酶活测定表明，AOS在黄皮小青豆（高抗）中的酶活约为在徐疃大豆（高感）中的4倍。该结果显示，GmAOS和GmAOS1基因在不同抗性材料中的表达量不同，斜纹夜蛾诱导时达到最大表达量所需要的时间不同，AOS在不同抗性材料中的酶活性不同，这些差异可能是黄皮小青豆（高抗）与徐疃大豆（高感）抗性不同的原因，AOS基因可能与植株的抗虫性有密切关系，利用GmAOS基因提高材料的抗虫性具有可行性。 为了鉴定GmAOS基因是否与材料的抗虫性有直接关系，利用过量表达GmAOS基因的方法得到了转基因烟草，Souther blot分析显示GmAOS基因以2~4个拷贝形式随机插入到烟草基因组中。昆虫生物学实验显示，在斜纹夜蛾对寄主的选择实验中，自由取食的20头斜纹夜蛾，喂养15小时后，有16头斜纹夜蛾选择了对照烟草，4头斜纹夜蛾选择了转基因烟草，且对照的叶面损失大于转基因叶面损失，该结果表明，斜纹夜蛾更喜欢啃噬对照叶片而避开转基因叶片；强迫饲喂实验中，转基因烟草喂养的斜纹夜蛾的相对增长率全部降低于对照，且对照烟草的叶面损失大于转基因烟草叶面损失。茸毛密度和形态分析显示，同一视野中，转基因烟草叶片上的平均茸毛数为60.2个，对照烟草叶片上的平均茸毛数为10.2个，即转基因烟草叶片上的平均茸毛数是对照烟草叶片上平均茸毛数的6倍，差异达到极显著水平；茸毛形态有明显差异，与对照茸毛相比，转基因烟草叶片上的茸毛要更加圆润、饱满、坚硬。酶学实验测定显示，与对照烟草相比，有90％转基因烟草提高了AOS酶活性；各有70％转基因烟草提高了POD和PPO酶活性；85％转基因烟草提高了CI酶活性。激素测定显示，与对照烟草相比，85％转基因烟草提高了内源茉莉酸含量；所有转基因烟草均提高了内源脱落酸的含量；有85％转基因烟草提高了赤霉素含量；转基因烟草提高了吲哚乙酸的含量也有一定提。以上结果表明GmAOS基因在提高大豆的抗虫性中起到了重要作用，可做为候选基因用于提高材料抗虫性进行抗虫育种。

Soybean (Glycine max (L). Merr) leaf-feeding insects affect the yield and quality of soybean. In this study, the 58 accessions were tested for the antibiosis to cotton worm (Prodenia litura, Fabricius) under inoculation condition according to the larval and pupal weight. The result indicated that both of the larval weight and pupal weight were all significantly different among varieties. Low larval and pupal weight indicates the high level of antibiosis of material. Larval weight was positively correlated with pupal weight. According to the results of 2004 and 2005, three genotypes, Huangpixiaoqingdou (HPXQD), shandongdadou and PI229358 were identified to be high resistant to cotton worm; Three genotypes, Xutuandadou (XTDD), mianyangbaimaodou and zhongyanghouzimao were identified to be high susceptible to cotton worm. These genotypes can be used to clone gene and functional analysis and inheritance study. GmAOS and its promoter was isolated from soybean use RACE. The full length sequence of GmAOS were 1789 bp which containing an ORF, encoding for 519 amino acids polypeptide with a calculated molecular mass of 58.3 kDa and an pI of 8.97. GmAOS had one exon and the protein possesses serine, threonine and tyrosine. The N-terminal region of GmAOS displayed features of a typical chloroplast targeting peptide. GmAOS has two copies in soybean and promoter of GmAOS contained two TATA-box, one CAAT-box, one GA inducing elements (TAACAA), one W-box element, two salt and pathogen inducing elements (GAAAAA) and one G-box (CACGTG) can be induced by JA . The different expression patterns of GmAOS and GmAOS1 (ABB917763) were analyzed in different organs and induced by various treatments in two soybean accessions (XTDD, HS and HPXQD, HR). GmAOS and GmAOS1 transcript levels correlated with materials resistance grades. GmAOS mRNA and GmAOS1 mRNA were widely distributed among all organs examined. GmAOS and GmAOS1 had higher transcript levels in HPXQD (HR) than that in XTDD (HS). GmAOS and GmAOS1 gene had responded to cool(4℃)/heat(45℃)/JA/SA/CuSO4<下标!>/Cut/ SMV treatments. Cotton worm showed a strong inducement of the GmAOS and GmAOS1 transcript. In XTDD (HS), GmAOS reached the maximum mRNA level after induced by 24 h in XTDD (HS) while after induced by 3 h in HPXQD (HR). GmAOS1 reached the maximum mRNA level after induced by 12 h in XTDD (HS) while after induced by 3 h in HPXQD (HR). The activity of AOS in HPXQD (HR) was 4 times than that of XTDD (HS). These results showed that GmAOS and GmAOS1 in XTDD (HS) reached the maximum mRNA level significantly later than that of HPXQD (HR). This result also showed that the GmAOS and GmAOS1 transcript levels were correlated with materials resistance grade to cotton worm and overexpression of GmAOS might be one way to improve defense abilities in higher plants. To further examine the effect of GmAOS gene, it was overexpressed in tobacco and genomic southern blot showed that it had 2~4 copy in transgenic tobacco. Insect bioassay was performed to test the insecticidal activity of the GmAOS overexpressed in transgenic tobacco plants. Host selection experiment showed after 15 h, 16 cotton worms had migrated controls, while 4 cotton worms chosed transgenic tobacco plants. The result indicated that cotton worms showed more preference to controls than transgenic tobacco, transgenic tobacco leaves were significantly less damaged compared to the control and the transgenic tobacco plant decreased the RGR of cotton worms. Scanning electron micrographs revealed that the average number of trichomes on transgenic tobacco plant leaves was 60.2 and the average number of trichomes on control tobacco plant leaves was 10.2 in the same field of vision. Analysis of enzyme activities showed that, in compared to the control, 90％ transgenic tobacco plants enhanced AOS activity, 70％ transgenic tobacco plants enhanced PPO and POD activity respectively and 85％ transgenic tobacco plants enhanced CI activity. The hormone assay indicated that, in contrast to the control, 85％ transgenic tobacco plants enhanced the contents of JA and GA respectively，all the transgenic tobacco plants enhanced the contents of ABA and the content of IAA also increased in transgenic tobacco. The results presented in this work contribute to reveal the importance of GmAOS in plant defense.and overexpression of GmAOS might be a useful tool for improving resistance against cotton worms in high plants.